Studies indicate that Mg2+ deficiency is an important etiological factor in many diseases and disorders, possibly including salivary gland dysfunction. A large proportion of the U.S. population, as many as 75%, may face a latent Mg2+ deficiency. Mg2+ plays a critical role in many cell functions, for example, Mg2+ forms complex substrates with ATP, citrate, etc.; Mg2+ is a co-factor for kinases, phosphatases, and synthetases; Mg2+ stabilizes the structure of membranes and ribosomes during protein synthesis; and Mg2+ regulates ion channel, exchanger, cotransporter, and pump activities. We found that Mg2+ effectively regulates the intracellular Na+ concentration and affects intracellular pH in rat sublingual acinar cells. Since intracellular pH and Na+ level are very important for regulating the fluid secretion process, Mg2+ may play a vital role in the modulation of the secretory function of exocrine salivary glands. However,the regulation of cell Mg2+ homeostasis is still poorly understood. In preliminary studies, we found that in rat sublingual acinar cells, the intracellular free Mg2+ concentration is regulated by the Mg2+ transport systems in the plasma membrane and by an intracellular Mg2+ pool. Mg2+ mobilization from the intracellular pool may depend on Ca2+, although the underlying mechanisms of Mg2+ transport in the plasma membrane and intracellular pool are still unknown. thus, the purpose of this project is to explore the Mg2+ uptake and efflux pathways in the plasma membrane and intracellular pool. 1) we will characterize the Mg2+ transport pathways in the plasma membrane using Mg2+-sensitive fluorescent indicator, mag-fura-2, to monitor Mg2+ influx and extrusion in isolated sublingual acini or single cells, and using cation channel blockers, activators, and ion exchange inhibitors to characterize the Mg2_ influx and efflux pathways. Also, we will apply the activators and inhibitors of protein kinase C and manipulate intracellular cAMP levels to clarify the regulatory mechanism of Mg2+ fluxes. 2) we will explore the mechanisms of Mg2+ uptake and release in the intracellular Mg2+ pool, in particular the role of Ca2+ in Mg2+ movement. In this project, we will clarify the role of Ca2+ in Mg2+ uptake and release by manipulating the intracellular free Ca2+ concentration. The results from these studies will provide insight for elucidating the regulatory mechanism of saliva secretion by Mg2+ and in preventing and treating salivary gland dysfunctions associated with Mg2+ deficiency.